Repurposing Peptide Nanomaterials as Synthetic Biomolecular Condensates in Bacteria.

Peptide nanomaterials exhibit diverse applications in vitro, such as drug delivery. Here, we consider the utility of de novo peptide nanomaterials to organize biochemistry within the bacterial cytoplasm. Toward this goal, we discovered that ABC coiled-coil triblock peptides form gel-like biomolecular condensates with a csat of 10 µM in addition to their well-known hydrogel-forming capabilities. Expression of the coiled-coil triblock peptides in bacteria leads to cell pole accumulation via a nucleoid occlusion mechanism. We then provide a proof of principle that these synthetic biomolecular condensates could sequester clients at the cell pole. Finally, we demonstrate that triblock peptides and another biomolecular condensate, RNase E, phase-separate as distinct protein-rich assemblies in vitro and in vivo. These results reveal the potential of using peptide nanomaterials to divide the bacterial cytoplasm into distinct subcellular zones with future metabolic engineering and synthetic biology applications.

Furin cleavage of the SARS-CoV-2 spike is modulated by O-glycosylation.

The SARS-CoV-2 coronavirus responsible for the global pandemic contains a novel furin cleavage site in the spike protein (S) that increases viral infectivity and syncytia formation in cells. Here, we show that O-glycosylation near the furin cleavage site is mediated by members of the GALNT enzyme family, resulting in decreased furin cleavage and decreased syncytia formation. Moreover, we show that O-glycosylation is dependent on the novel proline at position 681 (P681). Mutations of P681 seen in the highly transmissible alpha and delta variants abrogate O-glycosylation, increase furin cleavage, and increase syncytia formation. Finally, we show that GALNT family members capable of glycosylating S are expressed in human respiratory cells that are targets for SARS-CoV-2 infection. Our results suggest that host O-glycosylation may influence viral infectivity/tropism by modulating furin cleavage of S and provide mechanistic insight into the role of the P681 mutations found in the highly transmissible alpha and delta variants.

Furin cleavage of the SARS-CoV-2 spike is modulated by O-glycosylation.

The SARS-CoV-2 coronavirus responsible for the global pandemic contains a unique furin cleavage site in the spike protein (S) that increases viral infectivity and syncytia formation. Here, we show that O-glycosylation near the furin cleavage site is mediated by specific members of the GALNT enzyme family and is dependent on the novel proline at position 681 (P681). We further demonstrate that O-glycosylation of S decreases furin cleavage. Finally, we show that GALNT family members capable of glycosylating S are expressed in human respiratory cells that are targets for SARS-CoV-2 infection. Our results suggest that O-glycosylation may influence viral infectivity/tropism by modulating furin cleavage of S and provide mechanistic insight into the potential role of P681 mutations in the recently identified, highly transmissible B.1.1.7 variant.

Improved online LC-MS/MS identification of O-glycosites by EThcD fragmentation, chemoenzymatic reaction, and SPE enrichment.

O-glycosylation is a highly diverse and complex form of protein post-translational modification. Mucin-type O-glycosylation is initiated by the transfer of N-acetyl-galactosamine (GalNAc) to the hydroxyl group of serine, threonine and tyrosine residues through catalysis by a family of glycosyltransferases, the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferases (E.C. 2.4.1.41) that are conserved across metazoans. In the last decade, structural characterization of glycosylation has substantially advanced due to the development of analytical methods and advances in mass spectrometry. However, O-glycosite mapping remains challenging since mucin-type O-glycans are densely packed, often protecting proteins from cleavage by proteases. Adding to the complexity is the fact that a given glycosite can be modified by different glycans, resulting in an array of glycoforms rising from one glycosite. In this study, we investigated conditions of solid phase extraction (SPE) enrichment, protease digestion, and Electron-transfer/Higher Energy Collision Dissociation (EThcD) fragmentation to optimize identification of O-glycosites in densely glycosylated proteins. Our results revealed that anion-exchange stationary phase is sufficient for glycopeptide enrichment; however, the use of a hydrophobic-containing sorbent was detrimental to the binding of polar-hydrophilic glycopeptides. Different proteases can be employed for enhancing coverage of O-glycosites, while derivatization of negatively charged amino acids or sialic acids would enhance the identification of a short O-glycopeptides. Using a longer than normal electron transfer dissociation (ETD) reaction time, we obtained enhanced coverage of peptide bonds that facilitated the localization of O-glycosites. O-glycosite mapping strategy via proteases, cut-off filtration and solid-phase chemoenzymatic processing. Glycopeptides are enriched by SPE column, followed by release of N-glycans, collection of higher MW O-glycopeptides via MW cut-off filter, O-glycopeptide release via O-protease, and finally detected by LC-MS/MS using EThcD.

Should 15° of valgus coronal-plane deformity be the upper limit for a total ankle arthroplasty?

AIMS: Preoperative talar valgus deformity ≥ 15° is considered a contraindication for total ankle arthroplasty (TAA). We compared operative procedures and clinical outcomes of TAA in patients with talar valgus deformity ≥ 15° and < 15°. METHODS: A matched cohort of patients similar for demographics and components used but differing in preoperative coronal-plane tibiotalar valgus deformity ≥ 15° (valgus, n = 50; 52% male, mean age 65.8 years (SD 10.3), mean body mass index (BMI) 29.4 (SD 5.2)) or < 15° (control, n = 50; 58% male, mean age 65.6 years (SD 9.8), mean BMI 28.7 (SD 4.2)), underwent TAA by one surgeon. Preoperative and postoperative radiographs, Ankle Osteoarthritis Scale (AOS) pain and disability and 36-item Short Form Health Survey (SF-36) version 2 scores were collected prospectively. Ancillary procedures, secondary procedures, and complications were recorded. RESULTS: At mean 5.1 years follow-up (SD 2.6) (valgus) and 6.6 years (SD 3.3) (controls), mean AOS scores decreased and SF-36 scores increased significantly in both groups. Improvements in scores were similar for both groups - AOS pain: valgus, mean 26.2 points (SD 24.2), controls, mean 22.3 points (SD 26.4); AOS disability: valgus, mean 41.2 points (SD 25.6); controls, mean 34.6 points (SD 24.3); and SF-36 PCS: valgus, mean 9.1 points (SD 14.1), controls, mean 7.4 points (SD 9.8). Valgus ankles underwent more ancillary procedures during TAA (40 (80%) vs 13 (26%)) and more secondary procedures postoperatively (18 (36%) vs 7 (14%)) than controls. Tibiotalar deformity improved significantly (p < 0.001) towards a normal weightbearing axis in valgus ankles. Three valgus and four control ankles required subsequent fusion, including two for deep infections (one in each group). CONCLUSION: Satisfactory mid-term results were achieved in patients with preoperative valgus malalignment ≥ 15°, but they required more adjunctive procedures during and after TAA. Valgus coronal-plane deformity ≥ 15° is not an absolute contraindication for TAA if associated deformities are addressed. Cite this article: Bone Joint J 2020;102-B(12):1689-1696.

Changing surgeons improves outcome of subsequent primary total joint arthroplasty in previously dissatisfied patients.

We assessed whether patients who were dissatisfied with their previous primary hip (THA) or knee (TKA) arthroplasty, done by another surgeon, would have continued dissatisfaction or would have significant improvements in outcome scores following their subsequent primary THA or TKA. The majority of reasons provided for switching surgeons and/or institutions related to dissatisfaction with some aspect of their surgical experience specifically involving the surgeon-patient interaction itself. All 12 THA and TKA patients noted that their subsequent arthroplasty had decreased their pain, improved their function and that they were satisfied with their result. All patients had a statistically significant improvement in their Harris Hip Score or Knee Society Score, WOMAC and SF-36 questionnaires. This study demonstrates that previous dissatisfaction with a THA or THA does not predispose to a suboptimal outcome following subsequent primary hip or knee arthroplasty.

Severe metal-induced osteolysis many years after unipolar hip endoprosthesis.

BACKGROUND: Modularity of the femoral head-neck junction provides increased intraoperative flexibility to the surgeon. Complications of this modularity include damage to the trunnion, with subsequent bone and/or soft tissue loss from adverse reactions to metal debris. CASE DESCRIPTION: We describe two cases of severe metal-induced osteolysis and soft tissue damage requiring revision 10 and 13 years following implantation of a unipolar endoprosthesis. Damage to the trunnion resulted in severe acetabular and trochanteric osteolysis and soft tissue loss requiring complex revision surgery. LITERATURE REVIEW: Several reports have shown the trunnion, the head-neck interface, and the neck-stem couple as the causes of this early failure secondary to metal ion release from mechanical fretting corrosion or from crevice corrosion at these modular interfaces. These reports have been in association with a total hip prosthesis rather than a unipolar endoprosthesis. Revision of a unipolar endoprosthesis is most commonly attributable to stem loosening or acetabular erosion from the large femoral head articulating on the host acetabular cartilage and not owing to failure of the trunnion. PURPOSES AND CLINICAL RELEVANCE: Trunnion damage resulting in a severe reaction to metal debris with acetabular osteolysis, erosion of the greater trochanter, and loss of the abductor mechanism can occur years after implantation of a cementless unipolar endoprosthesis. This raises questions regarding long-term safety of the modular interface of a contemporary cementless stem and a large-diameter unipolar head. We recommend long-term followup of patients with a unipolar endoprosthesis as early recognition and treatment are required to avoid a potentially complex revision.

A new procedure for tibial spine avulsion fracture fixation.

Several techniques have been described to repair tibial spine avulsion fractures. Most of these methods use either internal fixation with a screw or suture fixation over a tibial tunnel bone bridge. This article presents a new technique for the surgical management of tibial spine avulsion fractures. The technique involves the creation of a suture mattress to compress and reduce the tibial spine into its fracture bed. The advantages of this technique are that it creates four points of fixation, aids with reduction, and allows for compression of the tibial spine fragment anatomically in its fracture bed. Level of evidence V.

Hydrothermal descriptive chemistry and single crystal structure determination of cesium and rubidium thorium fluorides.

Two new cesium thorium fluorides and three new rubidium thorium fluorides have been synthesized hydrothermally and structurally characterized. The structures of two polymorphs of CsTh(3)F(13) are described in space group P6/mmm with a = 8.2608(14) and c = 8.6519(17) and space group Pmc2(1) with a = 8.1830(16), b = 7.5780(15), and c = 8.6244(17). The analogous orthorhombic compound RbTh(3)F(13), with a = 8.1805(16), b = 7.4378(15), and c = 8.6594(17) in space group Pmc2(1), is also reported. Two other rubidium thorium fluorides are also described: RbTh(2)F(9) crystallizes in the space group Pnma where a = 8.9101(18), b = 11.829(2), and c = 7.4048(15), and Rb(7)Th(6)F(31) crystallizes in the space group R3 where a = 15.609(2) and c = 10.823(2). Comparison of these materials was made on the basis of their structures and synthesis conditions. The formation of these species in hydrothermal fluids appears to be dependent upon the concentration of the alkali fluoride mineralizer solution and, thus, the ratio of alkali ions to thorium in the system.

Hydro-thermally synthesized α-Ba(2)P(2)O(7).

Single crystals of α-Ba(2)P(2)O(7), dibarium diphosphate, were obtained under hydro-thermal conditions. The structure belongs to the diphosphate A(2)P(2)O(7) series with A being an alkaline earth cation. α-Ba(2)P(2)O(7) crystallizes isotypically with α-Sr(2)P(2)O(7). All atomic sites have site symmetry m with the exception of two O atoms which reside on general positions. Both Ba(2+) cations are coordinated by nine terminal O atoms from eclipsed diphosphate P(2)O(7) anions to form a three-dimensional network throughout the structure.

K(3)(Sc(0.875)Nb(0.125))Nb(2)O(9)H(1.75): a new scandium niobate with a unique cage structure.

Potassium scandium niobate hydroxide, K(3)(Sc(0.875)Nb(0.125))Nb(2)O(9)H(1.75), is a new scandium niobate with a unique cage structure. The structure contains two non-equivalent K(+) sites (3m and 6m2 site symmetry), one disordered Sc(3+)/Nb(5+) site (3m site symmetry), one Nb(5+) site (3m site symmetry), two O(2-) sites (m and mm2 site symmetry) and one H(+) site (m site symmetry). Both scandium and niobium have octahedral environments, which combine to form cages around potassium. One K atom lies in a cube-like cage built of seven octahedra, while the other K atom is encapsulated by an eight-membered trigonal face-bicapped prism. The cages form sheets that extend along the ab plane.

Ba(2)Ti(2)Si(2)O(9)F(2), a new titanium silicate.

Dibarium dititanium difluoride dioxide heptaoxidodisilicate, Ba(2)Ti(2)Si(2)O(9)F(2), is a new edge-sharing titanate with a unique titanium silicate framework. All atoms in the structure are in general positions. Titanium oxyfluoride octahedra combine with silicon tetrahedra to form a double stacked chain, which is the base unit of the layered framework. The Ba atoms lie in channels that extend along the a axis.